A long range goal of this project is to find a way to predict the three- dimensional structure of proteins on the basis of the amino acid sequences. In recent years and this year, we have made several key findings which have unified all previous studies of protein folding in this Section. Two manuscripts containing a mass of data have been completed by A. Fisher and H. Taniuchi in this year. In the following the findings and principles written in these manuscripts are summarized. A large number of homologous and hybrid 2-fragment complex of 4 different types were prepared from appropriate heme and apofragments or apoproteins of horse, tuna, yeast iso-1 and Candida cytochromes c. The complexes were characterized for structure, stability and Gibbs energy change for binding. The results have allowed us to assign 4 folding units which collectively represents cytochrome c folding. Unit 1 folds by itself and consists essentially of the right channel hydrophobic core and a part of heme. R.E. Dickerson and colleagues have found that the COOH-terminal helix contacts with the NH2-terminal helix to form a right channel. Units 2, 3 and 4, respectively are assigned to the cores on the left and right sides and at the bottom of heme. Based on evidence of the folding unit - unit interaction we propose that eukaryotic cytochrome c folds in two alternative ways depending on the temperature after attachment of heme to apoprotein by action of cytochrome c synthetase: Unit 1->3->2->4 or 1->2->3->4. Gibbs energy change for binding was found to vary depending on which species of heme fragment binds with which species of apofragment or apoprotein. This is more pronounced going from the less ordered complex to the more ordered. Evidence suggests that such Gibbs energy change difference of complexes containing the Fe-S bond is mainly due to perturbation of the folding unit-unit interaction, which may be mediated by contacting core groups. Based on this, a self- consistent set of residue substitutions to account for this Gibbs energy difference is assigned. Analysis of Gibbs energy change difference based on this assignment has suggested that a network of core group interaction may expand throughout the folding unit cores from Unit 1 as folding progresses. A model of this extra core interaction, core loop interaction/core loop coalescence which drives folding is proposed.